Role of

Transportation in
Spread of Porcine
Epidemic Diarrhea
Virus Infection,
United States
James Lowe, Phillip Gauger, Karen Harmon,
Jianqiang Zhang, Joseph Connor, Paul Yeske,
Timothy Loula, Ian Levis, Luc Dufresne,
and Rodger Main
After porcine epidemic diarrhea virus (PEDV) was
detected in the United States in 2013, we tested environ-
mental samples from trailers in which pigs had been trans-
ported. PEDV was found in 5.2% of trailers not contami-
nated at arrival, , suggesting that the transport process is a
source of transmission if adequate hygiene measures are
not implemented.
P
orcine epidemic diarrhea virus (PEDV) was detected
in herds of pigs in the United States during April 2013
(1). PEDV is a member of the Cornaviridae family that
produces a malabsorptive diarrhea secondary to atrophy
of the small intestinal villi (2). Initial clinical cases were
detected in herds in Indiana and Iowa during May 2013.
The virus spread rapidly across large geographic regions;
218 cases of infection were identifed in 16 states during
the frst 9 weeks of the outbreak (3). Subsequent testing of
historical samples collected during the week of April 15,
2013 identifed the index herd in Ohio (3). Veterinarians
became concerned about the role that facilities where pigs
are harvested for processing into food and the transporta-
tion equipment used to move pigs from farms to those fa-
cilities were playing in the spread of PEDV. These concerns
were based on evidence that equipment used to transport
live pigs transmits another enteric coronavirus, transmissi-
ble gastroenteritis virus, between sites in the United States
(J.F. Lowe, unpub. data).
Pigs are commonly transported to harvest
facilities in vehicles that have not been cleaned and
disinfected between loads. Implementation of all inall
out sites, which are sites in which pigs are grown and
all pigs in a group are removed before arrival of the next
group, limits the spread of disease introduced by trans-
port vehicles. In many cases, the risks and associated
costs of disease introduced late in the growing period are
thought to be less than the cost of cleaning and disinfect-
ing vehicles. Transport vehicles are often shared by dif-
ferent pig owners, enabling the spread of disease across
large regions.
The Study
The objective of this study was to assess the risks that
harvest facilities and transport vehicles engendered in
promoting the initial outbreak of a novel disease organism
by estimating the incidence of trailer contamination with
PEDV during the unloading process at harvest facilities.
Environmental samples were collected from 575 livestock
trailers before and after pigs were unloaded into holding
pens, or lairages, at 6 harvest facilities (83102 trailers
per facility) located in the central United States. Samples
were collected during a period of 23 days at each facility
during June 1420, 2013. For each trailer, the following
information was collected: transport company and trail-
er identifcation, time of unloading, dock used, whether
the truck driver stepped on the dock, and whether facil-
ity personnel entered the trailer. Sample collection con-
sisted of rubbing a phosphate-buffered salinemoistened
pad (Swiffer, Procter & Gamble, Cincinnati, OH, USA)
over an 900 cm
2
area of the trailer foor, 15 cm from the
rear door. The pad was placed in a sterile bag (Whirl-Pac,
NASCO, Fort Atkinson, WI, USA) and the liquid was
collected by applying manual pressure. The liquid was
transferred to a sterile tube (14mL Falcon Tube, Fisher
Scientifc, Chicago, IL, USA), immediately placed on ice,
and maintained at 4C during transport to the Iowa State
University Veterinary diagnostic laboratory. New latex
gloves were worn for each sample collection to minimize
the risk for cross-contamination.
RNA extraction was performed with 100 mL of
each environmental sample by using the MagMAX Vi-
ral RNA Isolation Kit (Life Technologies, Carlsbad, CA,
USA) and a Kingfsher 96 instrument (Thermo Scientifc,
Waltham, MA, USA) and Kingfsher program AM_1836_
DW_HV_v3 provided by the manufacturer of the viral
extraction kits. Viral RNA was eluted into 90 L of buf-
fer. Real-time reverse transcription PCR (rRT-PCR) was
performed on nucleic acid extracts by using the Path-ID
Multiplex One-Step RT-PCR Kit (Life Technologies) ac-
cording to the manufacturers recommendations. Prim-
ers and probe targeting conserved regions of the PEDV
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872 Emerging Infectious Diseases www.cdc.gov/eid Vol. 20, No. 5, May 2014
Author affliations: University of Illinois, Urbana, Illinois, USA
(J. Lowe); Iowa State University, Ames, Iowa, USA (P. Gauger,
K Harmon, J. Zhang, R. Main); Carthage Veterinary Service,
Ltd., Carthage, Illinois, USA (J. Connor); Swine Vet Center, P.A.,
St. Peter, MN, USA (P. Yeske, T. Loula); Iowa Select Farms,
Iowa Falls, Iowa, USA (I. Levis); and Seaboard Foods, Kansas City,
Missouri, USA (L. Dufresne)
DOI: http://dx.doi.org/10.3201/eid2005.131628
Roles of Transportation in PEDV Infection
nucleocapsid protein gene were as described (4) with mod-
ifcations specifc to the sequence isolated in North Amer-
ica deposited in GenBank (accession no. KF272920).
The forward primer sequence was 5-CGCAAAGACT-
GAACCCACTAACCT-3, the reverse primer sequence
was 5-TTGCCTCTGTTGTTACTTGGAGAT-3, and
the probe sequence was 5-TGTTGCCATTACCAC-
GACTCCTGC-3. Sequences were labeled by using
the FAM/ZEN/3 Iowa Black detector (Integrated DNA
Technologies, Coralville, IA, USA). All rRT-PCR re-
actions were conducted on an ABI 7500 Fast (Applied
Biosystems, Foster City, CA, USA) and results analyzed
by system software. Samples were tested separately from
routine diagnostic samples in the laboratory to minimize
risks for cross-contamination.
Before unloading, 38 (6.6%) of the 575 trailers
were contaminated with PEDV. The proportion of con-
taminated trailers ranged from 2% to 14.6% among the
6 harvest facilities; the facility level median was 5.0%.
Of the remaining 537, 28 (5.2%) that were not contami-
nated at arrival were contaminated in the unloading pro-
cess (Table).
Of the 38 trailers that were contaminated on arrival,
environmental samples from 13 (34.2%) were negative for
PEDV after unloading. Environmental samples from these
13 trailers tended to have higher cycle threshold values
than those from the 25 trailers that were positive before and
after unloading: 32.3 versus 30.6, respectively. This result
suggests that the pigs transported to the harvest facility on
the 13 trailers may not have been shedding PEDV, but in-
stead, the trailers had been contaminated by previous loads
of pigs, so viral quantities in the trailer were low or at the
limit of detection.
Contamination during unloading occurred at a higher
rate if harvest facility staff entered the trailer (OR 4.15,
95% CI 1.2713.54) or if unloading occurred immedi-
ately after unloading another trailer that was found to be
contaminated (OR 3.35, 95% CI 1.229.18). Facilities in
which more PEDV was identifed in truck trailers on arrival
had a higher overall incidence of contamination. This was
measured by multiplying the prevalence of contamination
at arrival by the inverse of the cycle threshold value from
trailers contaminated at arrival (R
2
= 0.32, p = 0.01). All
drivers stepped into the harvest facility at least once, lead-
ing to a high rate of contact between drivers, the trailers,
and the harvest facility.
Conclusions
Harvest facilities serve as a source of contact between
many swine farms with different health statuses. This
study suggests that collection points, such as harvest fa-
cilities and livestock auction markets, can be an effcient
source of contamination of transport vehicles that return to
pig farms and likely played a role in rapidly disseminating
PEDV across vast geographic regions shortly after PEDV
was frst identifed in the United States. These data also
suggest that the contamination of transport vehicles leaving
the harvest facilities increased as the prevalence of PEDV
positive transport vehicles and virus load coming into the
facility increased.
The results of this study suggest that proactive dis-
ease control measures should include improved sanitation,
hygiene, and segregation practices at collection points to
limit the spread of the agent early in the outbreak. Current
data suggest that novel agents, such as PEDV, may be
present in a country but remain undetected for an extend-
ed period. Thus, control measures may be implemented
too late to limit the spread of the disease through fomites
that are identifed, such as, in this instance, contaminated
vehicles returning from swine collection points. Simple
measures such as limiting contact between drivers and the
collection point and requiring drivers to remain on trucks
and out of the collection point during the unloading pro-
cess may have a dramatic effect on limiting the trans-
mission of novel agents. These biosecurity measures are
simple but require a coordinated effort between produc-
ers, transporters, harvest facility owners, and regulators
to achieve effective implementation. This study of PEDV
transmission by fomites should serve as an example of
the risks that a modern, highly technical animal protein
industry may encounter during a novel disease introduc-
tion. PEDVs introduction and subsequent spread in the
United States should spur action to minimize these risks
before a disease that can affect international trade or food
safety is introduced.
The National Pork Board, the National Pork Producers
Council, the American Association of Swine Veterinarians, and
the American Association of Swine Veterinarians Foundation
funded this project.
Dr Lowe is a clinical instructor in the Department of Veteri-
nary Clinical Medicine, University of Illinois, and is an adjunct
faculty member of Kansas State University and the University of
Minnesota. He also owns Lowe Consulting Ltd. and Production
Animal Consultation, LLC. His research interests include the ef-
fects of management on hostpathogen interactions and livestock
growth performance.
Emerging Infectious Diseases www.cdc.gov/eid Vol. 20, No. 5, May 2014 873

Table. Status of environmental samples from pig transport trailers
during an outbreak of porcine epidemic diarrhea virus infection,
Midwestern United States, June 2013*
PCR status
after unloading
PCR test status before unloading
Positive Negative Total
Positive 25 (4.3) 28 (4.9) 53 (9.2)
Negative 13 (2.3) 509 (88.5) 522 (90.8)
Total 38 (6.6) 537 (93.4) 575
*Values represent the number of trailers (% total) in each group. Samples
were gathered from 6 pig harvest facilities.

References
1. Stevenson GW, Hoang H, Schwartz KJ, Burrough ER, Sun D,
Madson D, et al. Emergence of porcine epidemic diarrhea
virus in the United States: clinical signs, lesions, and viral genomic
sequences. J Vet Diagn Invest. 2013;25:64954. http://dx.doi.
org/10.1177/1040638713501675
2. Saif LJ, Pensaert MB, Sestak K, Yeo S, and Jung K. Coronaviruses. In:
Zimmerman J, Karriker LA, Ramirez A, Schwartz KJ, Stevenson GW,
editors. Diseases of swine. Ames (IA): Wiley & Sons; 2012. p. 50124.
3. US Department of Agriculture, Animal and Plant Health Inspection
Service, National Veterinary Services Laboratories. Porcine Epidemic
Diarrhea virus (PEDv) Testing Data from NAHLN Laboratories:
PED/PEDv_weekly_report [cited 2013 Jul 3]; http://www.aasv.
org/aasv%20website/Resources/Diseases/PED/PEDv_weekly_
report_070313.pdf.
4. Kim SH, Kim IJ, Pyo HM, Tark DS, Song JY, Hyun BH. Multiplex
real-time RT-PCR for the simultaneous detection and quantifcation
of transmissible gastroenteritis virus and porcine epidemic diarrhea
virus. J Virol Methods. 2007;146:1727. http://dx.doi.org/10.1016/
j.jviromet.2007.06.021
Address for correspondence: James F. Lowe, Department of Veterinary
Clinical Medicine, 270 LAC, 1008 W Hazelwood Dr, Urbana, IL 61802,
USA; email: jlowe@illinois.edu
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874 Emerging Infectious Diseases www.cdc.gov/eid Vol. 20, No. 5, May 2014